The cerebellum is one of the brain areas involved in learning and memory formation. Long term synaptic plasticity is thought to play a pivotal role in supporting these functions. Moreover Immediate Early Genes (IEGs) expression and de novo protein synthesis and modification have been strictly associated with both LTP maintenance and memory consolidation and storage. We used rat parasagittal cerebellar slices as a model system in which specific patterns of stimulation delivered to the mossy fibers can induce both Long-Term Potentiation (LTP) and Long-Term Depression (LTD), depending on local inhibition and other regulating factors. Using Voltage Sensitive Dye (VSD) imaging we obtained high resolution maps of the spatial distribution of LTP/LTD induced from a Teta Burst Stimulus (TBS) application. Control and stimulated slices were then fixed at different times from the TBS application and processed for in situ hybridization or immunohystochemistry in order to detect IEGs mRNA expression patterns and protein modifications. We started analyzing CREB phosphorilation and c-fos expression. Two highly conserved signalling cascades seem to be involved in E-LTP to L-LTP conversion, PKA and MAPK, and both these pathway can activate CREB through phosphorilation. P-CREB expression has been suggested to initiate the protein synthesis that leads to the induction of L-LTP. c-fos is known to be rapidly and transiently induced in the Nervous System by a variety of stimuli and is thought to be directly involved in processes of neuronal plasticity such as LTP and under condition of learning. We correlate the expression pattern of c-fos and CREB mRNA and their protein distribution and/or phosphorlation with the LTP/LTD maps generated by VSD imaging. Preliminary data indicate a significant increase of P-CREB in the granular layer suggesting that CREB phosphorilation is induced as early as 15 minutes post TBS application. In situ hybridization experiments indicate a good correlation between c-fos induction and LTP distribution at 120 minutes post TBS. A similar up-regulation of c-Fos protein expression in the granular layer was detected by immunohystochemistry. We are planning further experiments to confirm these data and to test our experimental system in the presence of drugs that could interfere with the transcription, translation or post translational protein regulation.

Immediate early genes regulation in rat cerebellar cortex during long-term synaptic plasticity induction.

POLIMENI, MARIAROSA;GANDOLFI, DANIELA;CERRI, SILVIA;TRITTO, SIMONA;BLANDINI, FABIO;D'ANGELO, EGIDIO UGO
2011-01-01

Abstract

The cerebellum is one of the brain areas involved in learning and memory formation. Long term synaptic plasticity is thought to play a pivotal role in supporting these functions. Moreover Immediate Early Genes (IEGs) expression and de novo protein synthesis and modification have been strictly associated with both LTP maintenance and memory consolidation and storage. We used rat parasagittal cerebellar slices as a model system in which specific patterns of stimulation delivered to the mossy fibers can induce both Long-Term Potentiation (LTP) and Long-Term Depression (LTD), depending on local inhibition and other regulating factors. Using Voltage Sensitive Dye (VSD) imaging we obtained high resolution maps of the spatial distribution of LTP/LTD induced from a Teta Burst Stimulus (TBS) application. Control and stimulated slices were then fixed at different times from the TBS application and processed for in situ hybridization or immunohystochemistry in order to detect IEGs mRNA expression patterns and protein modifications. We started analyzing CREB phosphorilation and c-fos expression. Two highly conserved signalling cascades seem to be involved in E-LTP to L-LTP conversion, PKA and MAPK, and both these pathway can activate CREB through phosphorilation. P-CREB expression has been suggested to initiate the protein synthesis that leads to the induction of L-LTP. c-fos is known to be rapidly and transiently induced in the Nervous System by a variety of stimuli and is thought to be directly involved in processes of neuronal plasticity such as LTP and under condition of learning. We correlate the expression pattern of c-fos and CREB mRNA and their protein distribution and/or phosphorlation with the LTP/LTD maps generated by VSD imaging. Preliminary data indicate a significant increase of P-CREB in the granular layer suggesting that CREB phosphorilation is induced as early as 15 minutes post TBS application. In situ hybridization experiments indicate a good correlation between c-fos induction and LTP distribution at 120 minutes post TBS. A similar up-regulation of c-Fos protein expression in the granular layer was detected by immunohystochemistry. We are planning further experiments to confirm these data and to test our experimental system in the presence of drugs that could interfere with the transcription, translation or post translational protein regulation.
2011
Italian Journal of Anatomy and Embryology
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11571/583696
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